Calcium channel blockers (CCBs) may produce profound myocardial depression. Glucagon antagonized verapamil-induced hypotension and bradycardia in rats; however, glucagon's ability to antagonize other CCBs is unexplored. This study determined: a) if glucagon reverses verapamil-induced depression by a direct cardiac effect, b) if myocardial depression induced by diltiazem and nifedipine (representing different classes of CCBs) is also reversed by glucagon, and c) the glucagon concentration needed to reverse myocardial depression. Isolated rat hearts were perfused at a constant flow rate in a Langendorff preparation. Developed pressure (dP), contractility (+dP/dtmax), relaxation (-dP/dtmax), heart rate, and coronary vascular resistance were recorded. A CCB (n = 6, each blocker) was infused until >50% depression of contractility was achieved. Glucagon was then simultaneously infused (perfusion concentration of 0.6–1.1 × 10−7 M), and repeat cardiac variables were recorded. In a separate group of 36 hearts, glucagon dose response was determined. After producing a >50% depression in dP/dtmax with 3 μmol of diltiazem, a single concentration of glucagon was infused simultaneously into each heart (perfusion concentrations between 10−6 and 10−9 M) and percent recovery of baseline function was determined. Glucagon restored baseline contractility and dP with all three CCBs. Complete reversal of diltiazem-induced myocardial depression occurs at glucagon concentrations ≥5 × 10−7 M. We conclude that a) glucagon directly reverses myocardial depression from three classes of CCBs at concentrations achieved in vivo, and b) glucagon may be useful in the treatment of CCB-induced myocardial toxicity.